Alveolar macrophages (AM) produce two chemotactic factors for neutrophils that may be important in inflammatory processes in the lungs, a low molecular weight lipid and a 10 kDa protein. In past studies, we have identified the low molecular weight lipid as LTB4, but the protein species has not been completely characterized. Identification of this protein and the study of the regulation of its production is important in understanding the mechanisms by which inflammatory cells are recruited to the lungs. Our GOALS in these studies are to isolate and purify the protein chemoattractant; to isolate a specific cDNA in order to identify the amino acid sequence of the entire protein; to determine the relative contribution of this protein to the neutrophil chemotactic activity produced by the AM; and to study the regulation of this protein in vitro and in vivo, using an animal model of localized lung inflammation. We have found that porcine and human AM produce large amounts of this 10 kDa protein when stimulated with E. coli lipopolysaccharide (LPS). We have succeeded in purifying it 200 fold in a series of steps including cation exchange HP. We will purify the protein to homogeneity and determine the N-terminal amino acid sequence. We will raise a specific polyconal antibody to the protein in rabbits, and use this antibody to screen a cDNA library made from LPS stimulated AM in order to isolate a clone containing the specific cDNA to this protein. From this cDNA,, we will predict the amino acid sequence of the protein, which will permit a definitive comparison with other known cytokines. The preparation of the specific antibody and the cDNA probe will provide tools for the study of the expression/secretion of this protein in AM from an animal model of localized inflammation in order to understand when this protein is produced in vivo. We will test several pharmacologic interventions, including pre- and post-treatment with corticosteroids, in order to determine whether these drugs block the expression of the protein in vivo and limit lung inflammatory reactions. These studies will help define the role of an important AM product in the genesis of lung inflammation and they will determine whether steps that limit the production of this protein can be useful in limiting some types of lung inflammation in vivo.